Pneumocystis carinii is a major opportunistic pathogen of immuno- compromised patients. Because P. carinii cannot be cultured, molecular approaches have been utilized to identify and characterize antigens of this organism. Recombinant antigens can then be used to examine host immune responses to P. carinii infections. We have an ongoing project to characterize the antigens of both rat and human P. carinii.. We have previously purified the major surface glycoprotein (MSG) of both rat and human pneu-mocystis using high-performance liquid chromatography. It is necessary to use P. carinii from both sources because antigenically they are different and specifically the major sur-face antigens in rat and human P. carinii are clearly different. Subsequently, we identified a number of clones from a cDNA library of rat P. carinii that contain genes encoding for the MSG. These clones are clearly related but not identical, demonstrating that multiple genes encode the MSG. We have continued studies to characterize potential antigens of P. carinii genes. We have cloned a number of human P. carinii MSG genes and have produced a recombinant fragment encoding a highly conserved region of human P. carinii MSGs that can be expressed at high levels. Preliminary studies have demonstrated that most humans have antibodies to the recombinant peptide, supporting the hypothesis that most humans have been previously infected with P. carinii. We are attempting to develop an ELISA using this recombinant fragment. We are also attempting to express other regions of human P. carinii MSG genes, and ultimately plan to use this panel of recombinant pep-tides to examine cellular and antibody responses to P. carinii in healthy and immunocom-promised patients. In addition, we have begun studies to characterize other P. carinii antigens. We have identified two families of genes related to the MSG whose expressions are regulated differently from MSG genes. We have also identified a family of genes encoding proteases related to the yeast kexin, which functions as a pro-protein processing enzyme. This protease appears to be expressed on the surface of P. carinii. The goal of this study is to better understand the pathogenesis of P. carinii pneumonia with the hope that we can use this information to control or prevent this disease.